ZR Fungal/Bacterial DNA MiniPrep Catalog No. D6005

Transcription

1 INSTRUCTION MANUAL ZR Fungal/Bacterial DNA MiniPrep Catalog No. D6005 Highlights Simple, efficient isolation of DNA (up to 25 µg/prep) from all types of tough-to-lyse fungi (e.g., yeast) and bacteria in as little as 15 minutes. State-of-the-art, ultra-high density BashingBeads are fracture resistant and chemically inert. Can be used with any bead mill, disrupter, or vortex that can accommodate standard 2 ml tubes. Omits the use of organic denaturants as well as proteinases. Contents Product Contents & Specifications..1 Product Description Protocol 3 Ordering Information..4 Xpedition Sample Prep Technologies..5 Related Products For Research Use Only Ver

2 Page 1 Satisfaction of all Zymo Research products is guaranteed. If you should be dissatisfied with this product please call The ZR BashingBead Lysis Tubes (0.5 mm) Cat # S , is still available for individual purchase. Product Contents ZR Fungal/Bacterial DNA MiniPrep (Kit Size) D6005 (50 preps.) Storage Temperature ZR BashingBead Lysis Tubes (0.1 & 0.5 mm) 1 50 Room Temp. Lysis Solution 40 ml Room Temp. Fungal/Bacterial DNA Binding Buffer ml Room Temp. DNA Pre-Wash Buffer 3 15 ml Room Temp. Fungal/Bacterial DNA Wash Buffer 50 ml Room Temp. DNA Elution Buffer 10 ml Room Temp. Zymo-Spin IV Spin Filters (Orange Tops) 50 Room Temp. Zymo-Spin IIC Columns 50 Room Temp. Collection Tubes 150 Room Temp. Instruction Manual 1 - Note - Integrity of kit components is guaranteed for up to one year from date of purchase. Reagents are routinely tested on a lot-to-lot basis to ensure they provide maximal performance and reliability. 2 For optimal performance, add beta-mercaptoethanol to 0.5%(v/v) i.e., 500 µl per 100 ml. 3 A precipitate may have formed in the DNA Pre-Wash Buffer during shipping. To completely resuspend the buffer, incubate the bottle at ºC for 30 minutes and mix by inversion. DO NOT MICROWAVE. Specifications Format Bead Beating, Spin Column Purification Sample Sources mg (wet weight) fungi or bacteria or up to 200 mg tissue. This equates to approximately 10 9 bacterial cells, 10 8 yeast cells and 10 7 mammalian cells. Spores, pollen, nematodes, as well as other microorganisms can also be sampled. DNA Purity High quality DNA is eluted with DNA Elution Buffer making it perfect for PCR (A 260/A 280 >1.8). Note - Trademarks of Zymo Research Corporation. This product is for research use only and should only be used by trained professionals. It is not intended for use in diagnostic procedures. Some reagents included with this kit are irritants. Wear protective gloves and eye protection. Follow the safety guidelines and rules enacted by your research institution or facility. Disruptor Genie is a trademark of Scientific Industries, Inc. and FastPrep is a registered trademark of Qbiogene, Inc. DNA Size Limits Capable of recovering genomic DNA up to and above 40 kb. In most instances, mitochondrial DNA and viral DNA (if present) will also be recovered. DNA Recovery Typically, up to 25 µg total DNA is eluted into 100 µl (35 µl minimum) DNA Elution Buffer per sample. Equipment Microcentrifuge, Vortex, Cell Disrupter/Pulverizer (recommended)

3 Page 2 Product Description The ZR Fungal/Bacterial DNA MiniPrep is designed for the simple, rapid isolation of DNA from tough-to-lyse fungi, including A. fumigatus, C. albicans, N. crassa, S. cerevisiae, S. pombe, as well as from mycelium and Gram (+) and (-) bacteria. The procedure is easy and can be completed in as little as 15 minutes: fungal and/or bacterial samples are added directly to a ZR BashingBead Lysis Tube (0.1 & 0.5 mm) and rapidly and efficiently lysed by bead beating (e.g., Xpedition Sample Processor, Page 5) without using organic denaturants or proteinases. The DNA is isolated and purified using our Zymo-Spin Technology and is ideal for downstream molecular-based applications including PCR, array, etc. A schematic of the ZR Fungal/Bacterial DNA MiniPrep procedure is shown below. Zymo Research offers the following for nucleic acid stabilization, DNA isolation, and RNA isolation. Zymo Research Supplier M Zymo Yeast Research Yeast Competitor M M Y Spores Y E. B coli B Spores Y Y E. B coli B 1) DNA/RNA Shield (R , R ) can be used to stabilize nucleic acids and inactivate infectious agents in a variety of samples, without the need for reagent removal. 2) Xpedition Fungal/ Bacterial DNA MiniPrep (D6206) incorporates DNA stabilization technology to stabilize fungal/bacterial samples collected in the field, as a part of the DNA isolation workflow. High-yield DNA is successfully isolated from Saccharomyces cerevisiae (spores) and Y E. coli and cells E. using coli the B ZR Fungal/Bacterial using the ZR DNA Fungal/Bacterial MiniPrep. DNA Equivalent Kit. amounts Equivalent of yeast amounts or bacteria of yeast were or processed bacteria were using processed the ZR Fungal/Bacterial using the ZR DNA Fungal/Bacterial MiniPrep or DNA the kit Kit from or the supplier kit from M. competitor Equal volumes M. Equal of eluted volumes DNA of eluted were DNA then analyzed were then in analyzed a 0.8% (w/v) in a agarose/ethidium 0.8% (w/v) agarose/ethidium bromide gel. The bromide size marker gel. The M size is a marker 1 kb ladder M is (Zymo a 1 kb Research). ladder (Zymo Research). 3) ZR Fungal/Bacterial RNA MiniPrep (R2014) utilizes BeadBashing technology, RT-PCR inhibitor removal, and Zymo-Spin Technology for isolation of ultra-pure total RNA from fungal/bacterial samples. For Technical Assistance, please contact those at Zymo Research s Technical Department at or to tech@zymoresearch.com.

4 Page 3 Protocol For optimal performance, add beta-mercaptoethanol (user supplied) to the Fungal/Bacterial DNA Binding Buffer to a final dilution of 0.5%(v/v) i.e., 500 µl per 100 ml. 1 This equates to approximately 10 9 bacterial cells, 10 8 yeast cells or 10 7 mammalian cells. 2 Cap tube tightly to prevent leakage. 1. Add mg (wet weight) fungal or bacterial cells 1 that have been resuspended in up to 200 µl of water or isotonic buffer (e.g., PBS) or up to 200 mg of tissue to a ZR BashingBead Lysis Tube (0.1 mm & 0.5 mm). Add 750 µl Lysis Solution to the tube Secure in a bead beater fitted with a 2 ml tube holder assembly and process at maximum speed for 5 minutes. Note: Required processing time will vary depending on the device and application and therefore should be evaluated on a case by case basis. For example, processing times may be as little as 3 minutes when using high-speed cell disrupters (e.g., the portable TerraLyzer Sample Processor, FastPrep -24, or similar) or as long as 20 minutes when using lower speeds (e.g., Disruptor Genie, or standard benchtop vortexes). See manufacturer s literature for operating information. 3. Centrifuge the ZR BashingBead Lysis Tube in a microcentrifuge at 10,000 x g for 1 minute. 4. Transfer up to 400 µl supernatant to a Zymo-Spin IV Spin Filter (Orange Top) in a Collection Tube and centrifuge at 7,000 x g for 1 minute. Note: Snap off the base of the Zymo-Spin IV Spin Filter (Orange Top) prior to use. 5. Add 1,200 µl of Fungal/Bacterial DNA Binding Buffer to the filtrate in the Collection Tube from Step 4. 3 The Zymo-Spin IIC Column has a maximum capacity of 800 µl. 6. Transfer 800 µl of the mixture from Step 5 to a Zymo-Spin IIC Column 3 in a Collection Tube and centrifuge at 10,000 x g for 1 minute. 7. Discard the flow through from the Collection Tube and repeat Step Add 200 µl DNA Pre-Wash Buffer to the Zymo-Spin IIC Column in a new Collection Tube and centrifuge at 10,000 x g for 1 minute. 9. Add 500 µl Fungal/Bacterial DNA Wash Buffer to the Zymo-Spin IIC Column and centrifuge at 10,000 x g for 1 minute. 10. Transfer the Zymo-Spin IIC Column to a clean 1.5 ml microcentrifuge tube and add 100 µl (35 µl minimum) DNA Elution Buffer directly to the column matrix. Centrifuge at 10,000 x g for 30 seconds to elute the DNA. Ultra-pure DNA is now ready for use in your experiments..

5 Page 4 Ordering Information Product Description Catalog No. Kit Size ZR Fungal/Bacterial DNA MicroPrep D preps. ZR Fungal/Bacterial DNA MiniPrep D preps. ZR Fungal/Bacterial DNA MidiPrep D preps. ZR-96 Fungal/Bacterial DNA Kit D6006 2x96 preps. For Individual Sale Catalog No. Amount Fungal/Bacterial DNA Binding Buffer D ml Lysis Solution D ml DNA Pre-Wash Buffer D ml Fungal/Bacterial DNA Wash Buffer D ml DNA Elution Buffer D ml ZR BashingBead Lysis Tubes (0.1 & 0.5 mm) 1 S Zymo-Spin IV Spin Filters (Orange Tops) C Zymo-Spin IIC Columns C Collection Tubes C C C ,000 1 The ZR BashingBead Lysis Tubes (0.5 mm) Cat # S , is still available for individual purchase. Lysis Instruments Description Cat. No. Amount Disruptor Genie, 120V w/ 2 ml tube holder assembly. Disruptor Genie, 230V w/ 2 ml tube holder assembly. TurboMix Attachment, 2 ml Permanently mounts to most existing Vortex Genie mixers converting them to a Disruptor Genie. S S S unit 1 unit 1 unit The Disruptor Genie with 2 ml tube holder from Scientific Industries, Inc. (Cat. No. S from Zymo Research Corp.)

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